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Structural/functional homology between the bacterial andeukaryotic cytoskeletonsLinda A Amos1, Fusinita van den Ent2 and Jan Lowe3

Structural proteins are now known to be as necessary for

controlling cell division and cell shape in prokaryotes as they are

in eukaryotes. Bacterial ParM and MreB not only have atomic

structures that resemble eukaryotic actin and form similar

filaments, but they are also equivalent in function: the assembly

of ParM drives intracellular motility and MreB maintains the

shape of the cell. FtsZ resembles tubulin in structure and in its

dynamic assembly, and is similarly controlled by accessory

proteins. Bacterial MinD and eukaryotic dynamin appear to have

similar functions in membrane control. In dividing eukaryotic

organelles of bacterial origin, bacterial and eukaryotic proteins

work together.

Addresses

MRC Laboratory of Molecular Biology, Hills Road, CambridgeCB2 2QH, UK1e-mail: [email protected]: [email protected]: [email protected]

Current Opinion in Cell Biology 2004, 16:18

This review comes from a themed issue on

Cell structure and dynamics

Edited by John A Cooper and Margaret A Titus

0955-0674/$ see front matter

2003 Elsevier Ltd. All rights reserved.

DOI 10.1016/j.ceb.2003.11.005

Abbreviations

AMPPNP 50-adenylyl-imidodiphosphate

EM electron microscopyGFP green fluorescent proteinGTP guanosine 50-triphosphateMAPs microtubule associated proteins

MD mitochondrion-dividing

PD plastid-dividingPH pleckstrin homology

Introduction

Almost all eukaryotes depend on polymers of actin andtubulin to organize their cytoplasm, although the same

basic elements are used in a variety of ways [1]. The

ancestors of these two self-assembling dynamic filamen-

tous proteins are believed to have evolved first in prokar-

yotes. The hypothesis that the bacteria cytoskeleton is

related to the eukaryote cytoskeleton was first established

when the bacterial Z-ring, which constricts the cell during

division, was visualized using fluorescence (Figures 1a,b)

and was shown to consistof FtsZ, a protein with a fold that

mirrors tubulin [2] and displays similar dynamic proper-

ties [3,4]. The relationship between eukaryotic and

prokaryotic filaments became pronounced when the pro-

teins MreB, Mbl and ParM [5,6,79] were shown to

possess structures (Figures 2 and 3) and dynamic proper-

ties [7,9] that resemble those of actin.

The ancestors of other eukaryotic cytoskeletal proteins are

likely to be identified in prokaryotes in the near future.

Owing to low sequence conservation, uncovering distant

relationships between structural proteins will depend on

comparisons of their macromolecular and atomic struc-tures, as well as their functional mechanisms; FtsZ amino

acid sequences show less than 20% identity to tubulin,

while MreB and ParM have 15% and 12% identity to actin,

respectively. Furthermore, prokaryotes seem to be more

diverse than eukaryotes because the only cell division

protein to be identified as common to almost all prokaryotic

organisms is FtsZ; however, other common elements

might have been missed because of sequence variation.

Determination of cell shapeActin filaments and microtubules usually cooperate in

controlling the shapes of eukaryotic cells [1]. Cell mem-

branes tend to be supported by an underlying layer of one

or other of these cytoskeletal filaments. Such a role foractin-like filaments seems to have originated in bacteria.

Two proteins, MreB and Mbl (MreB-like), each form

cables that follow a helical path close to the membrane ofBacillus subtilis (Figure 1d) [5,9]. Equivalent proteins

are found in most other non-spherical bacteria. Mutants

lacking MreB are round instead of rod-shaped; however,

some non-spherical bacteria that naturally lack actin-like

proteins control their shape by extending the cell-wall

only at the poles [10].

Investigation of the structure of MreB from Thermotoga

maritima by X-ray crystallography and electron micro-

scopy have amply confirmed that it is an actin homologue

[6]. In the crystal structure (Figure 3), the four subdo-

mains around the central nucleotide binding site in each

monomer show a strong similarity to those of actin. Moreimportantly, MreB was found to crystallize in such a way

that monomers were lined up to form filaments whose

longitudinal contacts confirm predicted contacts in an

actin filament. The 5.1 nm longitudinal spacing of sub-

units in MreB filaments, compared with 5.5 nm for F-actin, was confirmed in electron micrographs of sheets

and filaments that were assembled from purified protein

(Figure 2) MreB appears to form two-stranded filaments

(Figure 2c) similar to F-actin, except that the strands do

not twist around each other. The double-stranded fila-

ments further associate into pairs and larger bundles.

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MreB can form curved, as well as straight, bundles in vitro(Figure 2a), suggesting that a cooperative conformational

change can occur.

Chromosome separationIn eukaryotes, pairs of chromosomes are always moved

apart by a spindle that is constructed from tubulin-contain-

ing microtubules and is driven by microtubule-associated

motors. This is the most obvious feature that universally

differentiates eukaryotes from prokaryotes; however,

there are indications that bacteria do have simple cyto-

skeletal machines to assist in DNA segregation.

ParM self-assembly drives plasmid separation

The protein ParM [7] forms highly dynamic actin-like

filaments in the bacterium Escherichia coli and uses a

self-assembly mechanism for force production and trans-

port that is also used by eukaryotic actin. The crystal

structure of the ParM monomer [8] has a clear homol-

ogy to those of actin and MreB (Figure 3). A bonus result

from this study was a crystal structure of ParM without

bound nucleotide. Of the eukaryotic proteins, only the

Figure 1

(a) (b)

(c)

(d)

(e)

Light microscopy of fluorescently-labeled cytoskeletal filaments in

bacteria. (a) Visualization of the Z-ring in E. coli cells expressing GFP

FtsZ; two different views of a cell undergoing constriction. Reprinted

from [17], Copyright (1996) National Academy of Sciences, U.S.A. Bar

1 mm. (b) Brightfield and fluorescence images of an E. coli cell

expressing a low level of GFPFtsZ. Photobleaching experiments

revealed that the Z-ring exchanges subunits rapidly with a cytoplasmic

pool. Reprinted from [4], Copyright (2002) National Academy of

Sciences, U.S.A. Bar 2 mm. (c) Combined phase-contrast andimmunofluorescence microscopy of fixed E. coli cells expressing

wild-type levels of ParM. The fluorescent staining shows intracellular

actin-like filaments. Reprinted from [7], Copyright (2003), with

permission from OUP. (d) Localization of GFPMbl (MreB-like protein)

expressed in Bacillus subtilis shows dynamic helical bands.

Immunofluorescence microscopy of endogenous Mbl in fixed cells gave

similar images. Reprinted from [9], Copyright (2003), with permission

from Elsevier. Bar 4 mm. (e) E. coli cells expressing YFP (yellow

fluorescent protein)-MinD. After deconvolution, these optically sectioned

images of fixed cells reveal that MinD is present in helical filaments.

MinEGFP gave similar images. Reprinted from [47], Copyright (2003)

National Academy of Sciences, U.S.A.

Figure 2

(a)

(b) (c) (d)

Current Opinion in Cell Biology

Electron micrographs of negatively stained filaments. Both MreB [6] and

ParM [8] assemble into protofilaments with a subunit repeat close to

that in F-actin. MreB protofilaments associate into pairs but do not twist

to form helices like actin filaments; the 2-stranded filaments further

associate into straight or curved bundles (a-c); the various MreB

polymers all assemble together in the same conditions. (b) Individual

protofilaments, equivalent to those seen in atomic detail in the crystal

structure of MreB, can also assemble into flat sheets. (c) A pair of

double-stranded filaments; the inset is an enlarged filtered image. (d)

ParM double protofilaments have an even more remarkable structural

similarity to F-actin, as is clear from the enlarged filtered image (right).

Bars 100 nm.

2 Cell structure and dynamics

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actin-related protein ARP3 has been seen in the emptyform [11]. In both cases, the empty cleft is 258 wider

than in the ADP-filled form of the protein, indicating a

hinge-like movement between domains. Electron micro-

scopy of assembled filaments showed them to be double-

stranded and helical (Figure 2d), with a subunit arrange-

ment and longitudinal repeat similar to F-actin [8]. The

longitudinal repeat of 4.9 nm is slightly smaller than that

of MreB (5.1 nm) or actin (5.5 nm); as a result of this and

the fact that there are 12.5 monomers per repeat (as

opposed to 12.514 in F-actin), the distance between

crossovers is only 30 nm.

ParM is a product of one of the genes on the R1 plasmid,

which maintains its presence in the bacterial host by

segregating copies of itself towards the two cell poles,

before cell division. Plasmids of this type encode a

centromere-like DNA sequence, called parC in the case

of plasmid R1, and two proteins, called ParM and ParR.Duplicated plasmids are paired at their centromeric re-

gions through direct interactions with ParR. This ParR

parC complex interacts with ParM and stimulates its

ATPase activity. Immunofluorescence microscopy has

revealed ParM-containing filamentous bundles (Figure 1c)

along the longitudinal axis of E. coli[7].

Separation of bacterial chromosomes might be assisted in

a similar way; orthologues of some of the par genes have

been identified on chromosomal DNA (see Update).However, a DNA replication factory at the centre of the

cell [12] is thought to supply at least some of the force that

drives newly duplicated DNA towards opposite cell poles.

Segregation starts with the replication origins, which move

further from the centre as replication continues.

In eukaryotes, several types of protein complexes enable

actin assembly to drive the movement of membranous

organelles or to push forward the leading edge of the cell

membrane [13]. Formins [14] seem to represent the

closest system to the ParRparC complex, in that they

promote the assembly of a simple bundle of actin fila-ments. In such cases, assembly-driven activity might

follow the actoclampin model [15], in which each growing

filament is continuously tethered to a clamp molecule via

a site on the side of the endmost, ATP-bound monomer.

Addition of a new monomer on to the growing end triggersa cycle of ATP-hydrolysis and advancement of the clamp

molecule to the new subunit on the same filament. By

contrast, complexes that incorporate the actin-family pro-

teins ARP2/3 nucleate new forwards-pointing actin fila-

ments as branches from existing filaments [16]. The new

filaments must then make new contacts with the mem-

brane that is being pushed, using new ARP2/3-containingcomplexes.

Although MreB and Mbl helical bundles are also highly

dynamic in vivo [9], it is not known whether their

assembly can drive movement of other objects, in the

manner of ParM or actin. Nor is it known whether any ofthe bacterial actin-like filaments can serve as tracks for

motor protein-driven movement. As yet there is no evi-

dence for linear motor proteins in prokaryotes, apart from

those that move DNA [12].

Cytokinesis/septationConstriction and abscission in different cell types

The division of most cells involves constriction of the

membrane until there is a narrow connection between

the daughter cells (Figure 4) and this is followed by a

separate abscission step. One common theme in both

Figure 3


Ribbon structures of actin-family monomers: Actin.ADP, from pdb 1J6Z;

eukaryotic actin-related protein ARP3, from pdb 1K8K [11]; bacterial

ParM.ADP, from pdb 1MWM; ParM empty, from pdb 1MWK [8];

bacterial MreB.AMPPNP, from pdb 1JCG [6]. Each monomer has two

similar domains with a nucleotide binding site in the cleft between them.

The four subdomains (1A, 1B, 2A, 2B) in each protein are coloured blue,

yellow red and green. The crystal structure of ParM with and without

ADP [8] indicates a domain movement of 258, close to the

conformational change predicted for actin from a comparison of

actin.ADP with ARP3 in the empty state and from EM studies of F-actin

under different conditions [67]. The protofilament in the MreB crystal

structure provided the first atomic-resolution view of inter-subunit

contacts in an actin-family filament [6].

Structural/functional homology between the bacterial and eukaryotic cytoskeletons Amos, van den Ent and Lowe 3


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steps seems to be the use of filamentous GTP-binding

proteins (FtsZ, septins, dynamin), although they are notclosely related.

In bacteria, the Z-ring that is assembled from FtsZ

initially defines the site of the division plane [1719]

and is then actively involved in the constriction mechan-

ism. Other proteins are recruited to the ring, depending

on species. In E. coli and B. subtilis, these include FtsA,

another member of the actin family, and ZipA. In the

early stage of constriction, either ZipA or FtsA can sta-

bilize the Z-ring but FtsA is required for the final abscis-

sion [20,21]. Several other proteins are required to

coordinate changes in the cell membrane with formation

of a septum in the cell wall [18].

In fungi and animal cells, but apparently not in plants, the

division plane is first defined by a ring assembled from

septin filaments [22,23]. Myosin is then recruited [24] andactin bundles are assembled, organized by formins [14].

Acto-myosin sliding-filament contraction constricts the

cell diameter until it is filled by the central-spindle

bundle of microtubules (Figure 4).

In higher plants, the division plane is first defined by a

dynamic ring of microtubules [25] but these do not cause

constriction. Subsequently, other microtubules assemble

at right angles to the division plane, forming a phragmo-

plast [26], which is similar to an extended central spindle.The final stage of cytokinesis is, perhaps, most clear in

higher plants; Golgi-derived vesicles are transported

along the microtubules of the phragmoplast to the plane

of division, where they fuse with one another to form the

incipient cell wall. Members of the dynamin family are

essential components here [27] and also in animal cells

[28]; in this situation, the role of dynamin could be either

to package new membrane into tubes that can be deliv-

ered to the new cell wall and/or to remodel the newly

fused membrane.

Dynamic assembly of FtsZ and its control

FtsZ and tubulin both assemble into protofilaments with

a subunit spacing of 4.04.2 nm [29]. GTP, bound to a

pocket at the top of one subunit, comes into contact with

the bottom of the next subunit where a loop, known as

T7, on the latter surface is responsible for triggering GTPhydrolysis [30]. In the case of tubulin heterodimers, the

T7 loop ofa-tubulin is thought to hydrolyse GTP that is

bound to the b-tubulin in another dimer, but the T7 loop

ofb-tubulin simply traps the GTP that is bound to its own

a-tubulin partner. FtsZ might form homodimers as inter-

mediate complexes during assembly [31] but GTP is,

presumably, hydrolysed in every monomer. Both tubulinand FtsZ polymers become unstable after GTP hydro-

lysis and exhibit dynamic instability. In vivo, the Z-ring is

even more dynamic than microtubules [4]. The rapid

turnover of GTP means that it is difficult to study the

assembly of purified FtsZ in vitro, although some progress

has been made recently [3234]. However, the preciseform of the in vivo polymer remains unknown. It is

unlikely to closely resemble eukaryotic microtubules

because lateral contacts between tubulin protofilaments

are made by polypeptide loops that are missing from FtsZ

[2]. It might consist of just a pair of protofilaments [29,33].

Just as microtubule stability is controlled by microtubule

associated proteins (MAPs), assembly of FtsZ is pro-

moted in vivo by proteins such as FtsA, ZipA [1921]

and ZapA [35]. ZipA, which has a long unfolded P/Q rich

domain [36], perhaps best resembles typical MAPs, which

Figure 4

SeptinsMyosin IIFormins

Actin

AnillinSpectrin

DynaminSyntaxin

MTsMTs

FtsZFtsAZipAZapAMreB MreB

ParM ParM

MinCDMinCD

FtsB, FtsIFtsK, FtsLFtsN, FtsQ

FtsW

(a)

(b)


Some of the cytoskeleton proteins involved in cytokinesis. (a)

Representation of a section through an animal cell when the actin-

containing cleavage ring has contracted around the central bundle of

microtubules (MTs) of the extended mitotic spindle, after the

chromosomes have separated. Several proteins aside from actin are

required to form the cleavage ring and produce constriction. Dynamin

and syntaxin are required for membrane remodeling during the final

stage of separation. (b) Representation of a section through a dividing

bacterium. Proteins FtsA, ZipA and ZapA join the tubulin homologue

FtsZ in the Z-ring, which constricts the cell membrane during division.

Mutations in other Fts (filamenting temperature-sensitive) proteins

prevent invagination of the outer cell wall. The actin homologue, ParM,

separates copies of the plasmid on which it is encoded. Actin-like MreB

is needed to give the cells the correct shape and also to segregate the

chromosomes correctly. The Min proteins are needed to prevent a Z-ring

from forming too close to a pole.



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are unstructured in the absence of tubulin. MinC appears

to inhibit the assembly of unwanted Z-rings (discussed in

the next section) by binding to FtsZ polymer (and dis-

placing FtsA [31]). The structure of the MinC dimer is

compatible with a model in which it binds to the sides of

two adjacent subunits in a protofilament [37], possiblyinhibiting a lateral association of protofilaments that

might be required to form the Z-ring [38]. Alternatively,

MinC might cause disassembly by inducing curvature in

FtsZ protofilaments. There is a precedent for this second

mechanism, in the way that stathmin interacts with two

tubulin dimers [39]. However, there is no evidence that

MinC sequesters FtsZ in the way that stathmin seques-

ters tubulin. When E. coli or B. subtilis are under stress,

FtsZ is sequestered by a protein called SulA, which

prevents GTP hydrolysis by binding to the T7 loop[40], or YneA [41], which may perform a similar role.

The Min systems in bacteria

A protein complex, called MinCD, is thought to be

responsible for inhibiting FtsZ from assembling prema-

turely on to sites that will ultimately be at the centres of

the two future daughter cells. MinD binds cooperatively

to membranes in the presence of ATP and recruits MinC,

which is required for the regulation of Z-ring assembly

[31,37]. In B. subtilis, a protein called DivIVA recruits

MinCD to each polar region [42], leaving only the site at

the centre of the parent cell free to assemble a Z-ring but,in E. coli, MinCD is not localized in this way. Instead, a

soluble protein, MinE, prevents MinCD from inhibiting

Z-ring assembly at the central site [43,44,45,46]. The

activity of MinE sets up a surprising oscillation of all

the Min proteins. Originating from the cell centre, a ringof MinE progresses towards one pole, disassembling

MinCD as it moves. The depolymerized MinC and MinD

are then free to assemble near to the other pole. On

reaching the first pole, MinE is released to work on

MinCD polymers that are formed in the second half of

the cell. Each movement, from one half of the cell to the

other, takes 3050 s. Meanwhile, the whole cell graduallyincreases in length. Recent images with improved resolu-

tion [47] have shown that MinCD does not coat the

whole cell membrane near one pole but forms a long

winding filament (Figure 1e); its relationship to the MreB

filaments, if any, is still unclear.

MinD resembles dynamin

The similarities between dynamin and MinD might be

purely coincidental, but it is interesting to compare them

because they could share a common mechanism. The N-

terminal domain of dynamin is a regular GTPase, with a

strong similarity to Ras [48]. GTP regulates dimerization,

as well as interactions with membranes and with a variety

of actin-binding proteins [27,28]. The other domains

have important roles in these interactions [49]. A 3 D

reconstruction of a dynamin polymer surrounding a mem-

brane tubule has been calculated from electron micro-

scope images [50]. It shows a helical arrangement of

dimeric GTPase domains, which stand out from the

membrane on legs, consisting of the smaller, regulatory

domains. Contact with the membrane is via PH (pleck-

strin homology) domains at the ends of the legs.

The structure of MinD has been solved in the empty

state as well as when bound to the nonhydrolysable

ATP analogue AMPPNP (50-adenylyl-imidodiphosphate)

[5153]. It belongs to a group of ATPases that are related

to the P-loop GTPases [54]. The role of ATP is to

modulate the interaction of the protein with itself and

its partners, MinC, MinE and membranes [43,44,

45,46]. In vitro, MinD is known to dimerize with ATP

before assembling directly on to the membrane to form a

fairly tight helical polymer [43,44,45]. This polymer issuperficially reminiscent of the outer wall of the dynamin

helix, but presumably has no legs distancing it from the

membrane inside. Interaction with the membrane is via a

short C-terminal segment [55,56], which is much smaller

than the multidomain region of dynamin that follows the

GTPase domain.

One possibility is that the MinCD polymer that is

assembled in vivo (Figure 1) contains a tubule of fresh

membrane that becomes incorporated into the cell mem-

brane when released by MinE; such a mechanism would

help ensure an even insertion of new lipid along thelength of the cell. This would be similar to the proposed

role for dynamin in organizing membrane insertion during

the abscission of eukaryotic cells [27,28].

Division of chloroplasts and mitochondriaFtsZ, MinD and MinE are found in eukaryotes in

chloroplasts (derived from cyanobacteria) [57] and in

some primitive mitochondria (derived from a-proteobac-

teria) and they serve important functions in organelle

division [58,59]. FtsZ is presumably required for mark-

ing the division site and/or constricting the inner mem-

brane. Eukaryotic proteins, such as actin, organelle-specific dynamins [6061,62] and septins [63], are also

necessary for organelle division. Thus, the division

mechanism is a hybrid of prokaryotic constriction and

eukaryotic abscission. Most mitochondria lack FtsZ and,

apparently, use only eukaryotic division mechanisms.

Electron microscopy of chloroplast structure [64] shows

electron-dense plastid-dividing (PD) rings, on the cyto-

plasmic face of the outer membrane and on the stromal

face of the inner membrane. Mitochondria have a similar

structure, known as the mitochondrion-dividing (MD)

ring [65]. The chloroplast FtsZ ring localizes on the

stromal side of the inner PD ring. Both the Z-ring and

then the PD rings appear before division starts but the Z-

ring disappears before the final abscission takes place. An

attempt to isolate the PD rings in detergent [64] yielded

insoluble 5 nm filaments of unknown composition (but



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the absence of septins from plants might be worth rein-

vestigating [63]). A chloroplast-specific dynamin associ-

ates with the chloroplast outer membrane at a late stage of

division [60,61] and it might be of significance that

this is when the outer membrane needs to increase in

area, even more than the inner membrane.

The activity of the Z-ring has been more clearly tracked

in chloroplasts than in any bacterium, although we cannot

be certain that the constriction mechanism, in conjunc-

tion with that of eukaryotic mechanisms, is truly repre-

sentative of that in bacteria.

ConclusionsCytoskeletal proteins that are related in both structure

and function are currently being identified in bacteria andeukaryotic cells. The actin-like and tubulin-like proteins

in bacteria are evolutionarily related to their eukaryotic

counterparts, having similar functions and 3D structures.

Bacteria also have accessory proteins that control FtsZ

assembly in the way that MAPs control tubulin assembly,

although there is no evidence that any are evolutionary

precursors of MAPs. As yet, no associated proteins for

bacterial actin have been found; the search for them will

be an important objective in the near future. Researchers

will also continue to look for prokaryotic motor proteins

that might use FtsZ or bacterial actin filaments as tracks.

The role of dynamins in the division of chloroplasts andmitochondria, and their role in eukaryotic cytokinesis, is

another exciting area for further research. Finally, the full

role of MinD and its accessory proteins in bacteria and

chloroplasts demands declarative investigation.

UpdateRecent immunofluorescence studies of helical MreB fila-

ments in E. coli [66] have demonstrated that chromo-

somes fail to segregate properly in the round cells formed

when MreB is missing or mutated. These results suggest

that there is a bacterial chromosome segregation mechan-

ism that is homologous to active plasmid separation [7].

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of special interest

of outstanding interest

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30 s. The rapid turnover correlates well with the high GTPase activity ofFtsZ in vitro. A mutant FtsZ, whose activity is much reduced, showed acorresponding reduction in turnover. Thus, the assembly dynamics ofFtsZ, like those of tubulin, are regulated by GTP hydrolysis. Interestingly,the mutant cells divided with a normal cell-cycle time.

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22. Longtine MS, Bi E: Regulation of septin organization andfunction in yeast. Trends Cell Biol 2003, 13:403-409.

23. Macara IG, Baldarelli R, Field CM, Glotzer M, Hayashi Y, Hsu SC,Kennedy MB, Kinoshita M, Longtine M, Low C et al.: Mammalianseptins nomenclature. Mol Biol Cell 2002, 13:4111-4113.

24. Shu S, Liu X, Korn ED: Dictyostelium and Acanthamoebamyosin II assembly domains go to the cleavage furrow of

Dictyostelium myosin II-null cells. Proc Natl Acad Sci USA 2003,100:6499-6504.

25. Dhonukshe P, Gadella TW: Alteration of microtubule dynamicinstability during preprophase band formation revealed byyellow fluorescent protein-CLIP170 microtubule plus-endlabeling. Plant Cell 2003, 15:597-611.

26. Smith LG: Plant cell division: building walls in the right places .Nat Rev Mol Cell Biol 2001, 2:33-39.

27.

Kang B-H, Busse JS, Bednarek SY: Members of the Arabidopsisdynamin-like gene family, ADL1, are essential for plantcytokinesis and polarized cell growth. Plant Cell 2003,15:899-913.

Members of the Arabidopsis dynamin-related protein family are shown tobe essential for polarized cell expansion and cytokinesis; mutants showdefects in cell plate assembly andcell wall formation. Thedistributionof afunctional green fluorescent fusion protein is dynamic and localized

asymmetrically on the plasma membrane.28.

Thompson HM, Skop AR, Euteneuer U, Meyer BJ, McNiven MA:The large GTPase dynamin associates with the spindlemidzone and is required for cytokinesis. Curr Biol 2002,12:2111-2117.

In both mammalian and Caenorhabditis elegans cells, dynamin localizesto the spindle midzone and is enriched in spindle midbody extracts. In C.elegans, dynamin also localizes to newly formed cleavage furrow mem-branes. Embryos from a temperature-sensitive mutant and dynaminRNAi-treated embryos were defective in the late stages of cytokinesis,suggesting that dynamin plays an essential role in the final separation ofdividing cells.

29. Lowe J, Amos LA: Tubulin-like protofilaments in Ca2R-inducedFtsZ sheets. EMBO J 1999, 18:2364-2371.

30.

Scheffers DJ, de Wit JG, den Blaauwen T, Driessen AJ: GTPhydrolysis of cell division protein FtsZ: evidence that theactive site is formed by the association of monomers.

Biochemistry 2002, 41:521-529.In mixtures of wild-type FtsZ and proteins with a mutation in the T7 loop(on the opposite side of the molecule from the GTP binding site), mostmutants were found to inhibit wild-typeGTP hydrolysis.This confirmsthata T7 loop is required to complete the active site.

31. Justice SS, Garcia-Lara J, Rothfield LI: Cell division inhibitorsSulA and MinC/MinDblock septum formation at different stepsin the assembly of the Escherichia coli division machinery.Mol Microbiol 2000, 37:410-423.

32. Gonzalez JM, Jimenez M, Velez M, Mingorance J, Andreu JM,Vicente M, Rivas G: Essential cell division protein FtsZassembles into one monomer-thick ribbons under conditionsresembling the crowded intracellular environment. J Biol Chem2003, 278:37664-37671.

33. Oliva MA, Huecas S, Palacios JM, Martin-Benito J, Valpuesta JM,Andreu JM: Assembly of archaeal cell division protein FtsZ anda GTPase-inactive mutant into double-stranded filaments.

J Biol Chem 2003, 278:33562-33570.

34. Caplan MR, Erickson HP: Apparent cooperative assembly of thebacterial cell division protein FtsZ demonstrated by isothermaltitration calorimetry. J Biol Chem 2003, 278:13784-13788.

35. Gueiros-Filho FJ, Losick R: A widely conserved bacterial celldivision protein that promotes assembly of the tubulin-likeprotein FtsZ. Genes Dev 2002, 16:2544-2556.

36. OhashiT, Hale CA,de Boer PAJ, Erickson HP: Structural evidencethat the P/Q domain of ZipA is an unstructured, flexible tetherbetween the membrane and the C- terminal FtsZ-bindingdomain. J Bacteriol 2002, 184:4313-4315.

37. Cordell SC, Anderson RE, Lowe J: Crystal structure of thebacterial cell division inhibitor MinC. EMBO J 2001,20:2454-2461.

38. JohnsonJE, Lackner LL,de Boer PAJ: Targeting of (D)MinC/MinDand (D)MinC/DicB complexes to septal rings in Escherichiacoli suggests a multistep mechanism for MinC-mediateddestruction of nascent FtsZ rings. J Bacteriol 2002,184:2951-2962.

39. Gigant B, Curmi PA, Martin-Barbey C, Charbaut E, Lachkar S,Lebeau L, Siavoshian S, Sobel A, Knossow M: The 4A- x-ray

structure of a tubulin:stathmin-like domain complex. Cell2000,102:809-816.

40.

Cordell SC, Robinson EJH, Lowe J: Crystal structure of the SOScell division inhibitor SulA and in complex with FtsZ.Proc Natl Acad Sci USA 2003, 100:7889-7894.

SulA is shown to inhibit FtsZ assembly by binding to the T7 loop surface,which forms part of an active site when FtsZ is assembled into proto-filaments. The conformation of FtsZ in the complex with SulA is the sameas in crystals of FtsZ alone.

41. Kawai Y, Moriya S, Ogasawara N: Identification of a protein,YneA, responsible for cell division suppression during the SOSresponse in Bacillus subtilis. Mol Microbiol 2003, 47:1113-1122.

42. Harry EJ, Lewis PJ: Early targeting of Min proteins to the cellpoles in germinated spores of Bacillus subtilis: evidence fordivision apparatus-independent recruitment of Min proteins tothe division site. Mol Microbiol 2003, 47:37-48.

43.

Suefuji K, Valluzzi R, RayChaudhuri D: Dynamic assembly, of

MinD into filament bundles modulated by ATP, phospholipids,and MinE. Proc Natl Acad Sci USA 2002, 99:16776-16781.

This paper, together with [32,33,34], confirms the importance of themembrane as a partner in interactionsbetween theMin proteins,Min C, Dand E. EM images of MinD filaments assembled in the presence of ATPare shown here.

44.

Hu Z, Gogol EP, Lutkenhaus J: Dynamic assembly of MinD onphospholipid vesicles regulated by ATP and MinE. Proc NatlAcad Sci USA 2002, 99:6761-6766.

Includes an impressive EM image of MinD bound to phospholipidvesiclesandassembled into a helical array that deformed thevesiclesinto tubes inthe presence of ATP. Addition of MinE stimulated MinDs ATPase andcaused its release from the vesicles.

45. Lackner LL, Raskin DM, de Boer PAJ: ATP-dependentinteractions between Escherichia coli Min proteins andthe phospholipid membrane in vitro. J Bacteriol 2003,185:735-749.

46.

Hu Z, Saez C, Lutkenhaus J: Recruitment of MinC, an inhibitor ofZ-ring formation, to the membrane in Escherichia coli: role ofMinD and MinE. J Bacteriol 2003, 185:196-203.

A more detailed study. In the absence of lipid, ATP causes MinD todimerize and form a rather unstable complex with MinC. However, MinDbinds well to lipid vesicles in the presence of ATP and then recruits MinC.If MinD release via ATP hydrolysis is prevented, MinE can displace MinCthat is bound to the MinD-lipid complex but MinC does not displacebound MinE.

47.

Shih Y-L, Le T, Rothfield L: Division site selection in Escherichiacoli involves dynamic redistribution of Min proteins withincoiled structures that extend between the two cell poles .Proc Natl Acad Sci USA 2003, 100:7865-7870.

Imagesoffluorescently-labeled proteins showthat the threeMin proteins,all known to oscillate between the two poles of E. coli, are dynamicallyassociated with helices that wind around the cytoplasmic side of the cellmembrane. The authors also show that the E. coli MreB protein formsextended helices similar to those previously reported for B. subtilis [7].

TheMreB andMinCDE coiledarrays do notappear identicalbut this pointneeds to be confirmed by co-labeling the proteins in the same cells.

48. Niemann HH, Knetsch MLW, Scherer A, Manstein DJ, Kull FJ:Crystal structure of a dynamin GTPase domain in bothnucleotide-free and GDP-bound forms. EMBO J 2001,20:5813-5821.

49. Hinshaw JE: Dynamin and its role in membrane fission.Annu Rev Cell Dev Biol 2000, 16:483-519.

50. Zhang P, Hinshaw JE: Three-dimensional reconstruction ofdynamin in the constricted state. Nat Cell Biol 2001, 3:922-927.

51. Hayashi I, Oyama T, Morikawa K: Structural and functionalstudies of MinD ATPase: implications for the molecularrecognition of the bacterial cell division apparatus. EMBO J2001, 20:1819-1828.



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52. Cordell SC, Lowe J: Crystal structure of the bacterial celldivision regulator MinD. FEBS Lett 2001, 492:160-165.

53. Sakai N, Itou H, Watanabe N, Yao M, Tanaka I: The MinD proteinfrom the hyperthermophilic archaeon Pyrococcus horikoshii:crystallization and preliminary X-ray analysis.Acta Crystallogr DBiol Crystallogr 2001, 57:896-897.

54. Leipe DD, Wolf YI, Koonin EV, Aravind L: Classification andevolution of P-loop GTPases and related ATPases.J Mol Biol 2002, 317:41-72.

55. Szeto TH, Rowland SL, Rothfield LI, King GF: Membranelocalization of MinD is mediated by a C-terminal motif that isconserved across eubacteria, archaea, and chloroplasts.Proc Natl Acad Sci USA 2002, 99:15693-15698.

56. Hu ZL, Lutkenhaus J: A conserved sequence at the C-terminusof MinD is required for binding to the membrane andtargeting MinC to the septum. Mol Microbiol 2003,47:345-355.

57. Martin W, Rujan T, Richly E, Hansen A, Cornelsen S, Lins T,Leister D, Stoebe B, Hasegawa M, Penny D: Evolutionary analysisof Arabidopsis, cyanobacterial, and chloroplast genomesreveals plastid phylogeny and thousands of cyanobacterialgenes in the nucleus. Proc Natl Acad Sci USA 2002,99:12246-12251.

58. Vitha S, McAndrew RS, Osteryoung KW: FtsZ ring formationat the chloroplast division site in plants. J Cell Biol 2001,153:111-119.

59.

Reddy MSS, Dinkins R, Collins GB: Overexpression of theArabidopsis thaliana MinE1 bacterial division inhibitorhomologue gene alters chloroplast size and morphology intransgenic Arabidopsis and tobacco plants. Planta 2002,215:167-176.

Confocal and electron-microscopy of plants overexpressing MinE1revealed chloroplasts with abnormal shapes and sizes. This indicatesthat this protein probably has a similar role to bacterial MinE.

60.

Gao H, Kadirjan-Kalbach D, Froehlich JE, Osteryoung KW: ARC5,a cytosolic dynamin-like protein from plants, is part of thechloroplast division machinery. Proc Natl Acad Sci U S A 2003,100:4328-4333.

Unlike most mitochondria, chloroplasts have retained FtsZ for division.This paper (and [61,62]) show that a requirement for a dynamin-related

protein, with no obvious counterparts in prokaryotes, is common to bothtypes of endosymbiotic organelle. ARC5 is related to a group of dynamin-like proteins that are unique to plants. A GFPARC5 fusion proteinlocalizes to a ring at the chloroplast division site and ARC5 mutationscause enlarged, dumbbell-shaped chloroplasts. Import and protease

protection assays indicate that the ARC5 ring is positioned on the outersurface of the chloroplast.

61.

Miyagishima SY, Nishida K, Mori T, Matsuzaki M, Higashiyama T,Kuroiwa H, Kuroiwa T: A plant-specific dynamin-related proteinforms a ring at the chloroplast division site . Plant Cell 2003,15:655-665.

Analysis of synchronized chloroplasts showed the time at which a

dynamin-like protein is recruited to chloroplasts, forming a ring on thecytosolic side of the division site in the late stage of division. The ringconstricts until division is complete and then disappears.

62.

Nishida K, Takahara M, Miyagishima S, Kuroiwa H, Matsuzaki M,Kuroiwa T: Dynamic recruitment of dynamin for finalmitochondrial severance in a primitive red alga. Proc Natl AcadSci USA 2003, 100:2146-2151.

Previously, FtsZ was shown to localize to mitochondria in primitive algaeon the inside surface of the division site. Here, it is shown that FtsZ,dynamin andthe MD ring,an electron-dense structureon thecytoplasmicside of the mitochondrial membrane, act during different phases ofdivision. Before final separation, dynamin patches are recruited exter-nally, to the midpoint of the constricted MD ring.

63.

Takahashi S, Inatome R, Yamamura H, Yanagi S: Isolation andexpression of a novel mitochondrial septin that interacts withCRMP/CRAM in the developing neurones. Genes Cells 2003,8:81-93.

This presence of a septin in mitochondria might mean that septins are

candidates for components of the MD ring [60

] (and also chloroplast PDrings [64]).

64.

Kuroiwa H, Mori T, Takahara M, Miyagishima S, Kuroiwa T:Chloroplast division machinery as revealed byimmunofluorescence and electron microscopy. Planta 2002,215:185-190.

The behaviour of FtsZ and PD rings, both components of the chloroplastdivision machinery,was followed during the course of division.FtsZ formsa ring onthe stromal division site beforeinvagination starts. Then theinner(stromal) and outer (cytosolic) PD rings appear. The FtsZ ring remains atthe leading edge of the constriction but does not change width althoughthe volume of the outer PD ring gradually increases. The FtsZ ringdisappears before the final stage of chloroplast constriction.

65. Kuroiwa T, Kuroiwa H, Sakai A, Takahashi H, Toda K, Itoh R:The division apparatus of plastids and mitochondria. Int RevCytol 1998, 181:1-41.

66. Kruse T, Mller-Jensen J, Lbner-Olesen A, Gerdes K:

Dysfunctional MreB inhibits chromosome segregation inEscherichia coli. EMBO J 2003, 22:5283-5292.

67. Egelman EH:Actins prokaryotic homologs. Curr Opin Struct Biol2003, 13:244-248.



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